3H-DOB (4-bromo-2,5-dimethoxyphenylisopropylamine) labels a guanyl nucleotide-sensitive state of cortical 5-HT2 receptors

3H-(+/-)-4-Bromo-2,5-dimethoxyphenylisopropylamine (3H-DOB), a putative agonist radioligand, was synthesized and used to label 5-HT2 receptors in a particulate fraction prepared from rat frontal cortex tissue homogenates. The specific binding (defined by the difference in 3H-DOB binding in the presence and absence of 10(-6) M cinanserin, a potent and specific 5-HT2 antagonist) displayed high affinity (KD = 4.1 X 10(-10) M) and saturability with a Bmax of 17.9 fmol/mg of protein. The distribution of specific 3H-DOB binding in nine brain regions correlated closely with the distribution of 3H-ketanserin (an antagonist radioligand)-labeled 5-HT2 receptors. Competition studies in frontal cortex homogenates using a variety of compounds revealed a distinct 5-HT2 receptor pharmacology. A series of 5-HT2 antagonists exhibited high affinities in competition studies for specific 3H-DOB binding. The absolute potencies of these antagonists as well as their order of potencies closely correlated with their potencies in competing for 3H-ketanserin-labeled brain 5-HT2 receptors. A series of 5-HT2 agonists also exhibited high affinities in competition studies for specific 3H-DOB binding. Although the order of potencies of these agonists was similar to their order in competing for 3H-ketanserin-labeled brain 5-HT2 receptors, the agonists displayed 10-100-fold higher affinities for the 3H-DOB-labeled sites than for the 3H-ketanserin-labeled sites. The level of specific 3H-DOB binding in the frontal cortex homogenates was approximately 5% of the levels of 3H-ketanserin-labeled 5-HT2 receptors (358 fmol/mg of protein). Taken together, these results indicate that 3H-DOB labels a subset of brain 5-HT2 receptors that has high affinity for agonists as well as antagonists); 3H-ketanserin appears to label both subsets of brain 5-HT2 receptors. Antagonists apparently do not discriminate between these two subsets of 5-HT2 receptors. 3H-DOB specific binding to 5-HT2 receptors was potently inhibited by guanosine 5'-(beta, gamma-imido)triphosphate and guanosine 5'-O-(3-thio)triphosphate (nonhydrolyzable derivatives of GTP) with IC50 values of 42 and 21 nM, respectively, whereas adenosine 5'-(beta, gamma-imido)triphosphate and adenosine 5'-O-(3-thio)triphosphate (nonhydrolyzable derivatives of ATP) had no effect. In summary, 3H-DOB specific binding displays the pharmacological characteristics of a 5-HT2 receptor.